I am trying to set up an assay to study the internalization of an antibody into HEK cells. The antibody binds to a surface exposed receptor and is proposed to be internalized upon binding.
I have so far tried to incubate the cells with biotinylated antibody (which clearly binds to the cells) and thereafter stripp the cells from surface bound antibodies followed by cell lysis. I have however not been able to get a positive result.
I am unsure of if my assay set up is ok regarding amount of cells, lyzis buffer, stripping buffer etc.
I do not have any positive control either so it is difficult for me to detect where the problem is.

Does anyone have a good protocol of an internalization assay?
Or does anyone have any hints or tips to give me?

I assume you do western blot after lysis, correct? Choosing the right lysis buffer is very important in such experiments. My question is why don't you run immunofluorescence experiment with your cells? this way you can view where the antibody binds to. What is your protocol, I'll have a look at it.